Aim. Feasibility and reliability of tissue Doppler imaging-(TDI-) based elastography for cervical quantitative stiffness assessment during all three trimesters of pregnancy were evaluated. Materials and Methods. Prospective case-control study including seventy-four patients collected between the 12th and 42nd weeks of gestation. The tissue strain (TS) was measured by two independent operators as natural strain. Intra- and interoperator intraclass correlation coefficient (ICC) agreements were evaluated. Results. TS measurement was always feasible and exhibited a high performance in terms of reliability (intraoperator ICC-agreement = 0.93; interoperator ICC agreement = 0.89 and 0.93 for a single measurement and for the average of two measurements, resp.). Cervical TS showed also a significant correlation with gestational age, cervical length, and parity. Conclusions. TS measurement during pregnancy demonstrated high feasibility and reliability. Furthermore, TS significantly correlated with gestational age, cervical length, and parity.